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      Novel genomic islands and a new vanD-subtype in the first sporadic VanD-type vancomycin resistant enterococci in Norway

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          Abstract

          Background

          Vancomycin-resistant enterococci (VRE) represent several types of transferable vancomycin resistance gene clusters. The vanD type, associated with moderate to high level vancomycin resistance, has only sporadically been described in clinical isolates. The aim of this study was to perform a genetic characterization of the first VanD-type VRE strains detected in Norway.

          Methods

          The VanD-type VRE-strains (n = 6) from two patient cases were examined by antimicrobial susceptibility testing and whole genome sequencing (WGS) to uncover Van-phenotype, strain phylogeny, the vanD gene clusters, and their genetic surroundings. The putative transferability of vanD was examined by circularization PCR and filter mating.

          Results

          The VanD-type Enterococcus faecium (n = 4) and Enterococcus casseliflavus (n = 2) strains recovered from two cases (A and B), expressed moderate to high level vancomycin resistance (MIC 64—>256 mg/L) and various levels of teicoplanin susceptibility (MIC 2—>256 mg/L). WGS analyses revealed phylogenetically different E. faecium strains (A1, A2, and A3 of case A and B1 from case B) as well as vanD gene clusters located on different novel genomic islands (GIs). The E. casseliflavus strains (B2 and B3 of case B) were not clonally related, but harbored nearly identical novel GIs. The vanD cluster of case B strains represents a novel vanD-subtype. All the vanD-GIs were integrated at the same chromosomal site and contained genes consistent with a Clostridiales origin. Circular forms of the vanD-GIs were detected in all strains except B1. Transfer of vanD to an E. faecium recipient was unsuccessful.

          Conclusions

          We describe the first VanD-type E. casseliflavus strains, a novel vanD-subtype, and three novel vanD-GIs with a genetic content consistent with a Clostridiales order origin. Despite temporal occurrence, case A and B E. faecium strains were phylogenetically diverse and harbored different vanD subtypes and vanD-GIs.

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          Most cited references48

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          Trimmomatic: a flexible trimmer for Illumina sequence data

          Motivation: Although many next-generation sequencing (NGS) read preprocessing tools already existed, we could not find any tool or combination of tools that met our requirements in terms of flexibility, correct handling of paired-end data and high performance. We have developed Trimmomatic as a more flexible and efficient preprocessing tool, which could correctly handle paired-end data. Results: The value of NGS read preprocessing is demonstrated for both reference-based and reference-free tasks. Trimmomatic is shown to produce output that is at least competitive with, and in many cases superior to, that produced by other tools, in all scenarios tested. Availability and implementation: Trimmomatic is licensed under GPL V3. It is cross-platform (Java 1.5+ required) and available at http://www.usadellab.org/cms/index.php?page=trimmomatic Contact: usadel@bio1.rwth-aachen.de Supplementary information: Supplementary data are available at Bioinformatics online.
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            Basic local alignment search tool.

            A new approach to rapid sequence comparison, basic local alignment search tool (BLAST), directly approximates alignments that optimize a measure of local similarity, the maximal segment pair (MSP) score. Recent mathematical results on the stochastic properties of MSP scores allow an analysis of the performance of this method as well as the statistical significance of alignments it generates. The basic algorithm is simple and robust; it can be implemented in a number of ways and applied in a variety of contexts including straightforward DNA and protein sequence database searches, motif searches, gene identification searches, and in the analysis of multiple regions of similarity in long DNA sequences. In addition to its flexibility and tractability to mathematical analysis, BLAST is an order of magnitude faster than existing sequence comparison tools of comparable sensitivity.
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              QUAST: quality assessment tool for genome assemblies.

              Limitations of genome sequencing techniques have led to dozens of assembly algorithms, none of which is perfect. A number of methods for comparing assemblers have been developed, but none is yet a recognized benchmark. Further, most existing methods for comparing assemblies are only applicable to new assemblies of finished genomes; the problem of evaluating assemblies of previously unsequenced species has not been adequately considered. Here, we present QUAST-a quality assessment tool for evaluating and comparing genome assemblies. This tool improves on leading assembly comparison software with new ideas and quality metrics. QUAST can evaluate assemblies both with a reference genome, as well as without a reference. QUAST produces many reports, summary tables and plots to help scientists in their research and in their publications. In this study, we used QUAST to compare several genome assemblers on three datasets. QUAST tables and plots for all of them are available in the Supplementary Material, and interactive versions of these reports are on the QUAST website. http://bioinf.spbau.ru/quast . Supplementary data are available at Bioinformatics online.
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                Author and article information

                Contributors
                Role: Data curationRole: Formal analysisRole: MethodologyRole: SoftwareRole: ValidationRole: VisualizationRole: Writing – original draft
                Role: ConceptualizationRole: Data curationRole: Formal analysisRole: Funding acquisitionRole: MethodologyRole: SoftwareRole: SupervisionRole: ValidationRole: VisualizationRole: Writing – review & editing
                Role: ConceptualizationRole: Funding acquisitionRole: InvestigationRole: SupervisionRole: ValidationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: InvestigationRole: ValidationRole: Writing – original draft
                Role: Data curationRole: InvestigationRole: ValidationRole: Writing – original draft
                Role: ConceptualizationRole: Funding acquisitionRole: SupervisionRole: ValidationRole: Writing – review & editing
                Role: ConceptualizationRole: Data curationRole: Formal analysisRole: Funding acquisitionRole: InvestigationRole: MethodologyRole: Project administrationRole: SupervisionRole: ValidationRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS One
                plos
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                23 July 2021
                2021
                : 16
                : 7
                : e0255187
                Affiliations
                [1 ] Research Group for Host-Microbe Interactions, Department of Medical Biology, University of Tromsø (UiT) – The Arctic University of Norway, Tromsø, Norway
                [2 ] Norwegian National Advisory Unit on Detection of Antimicrobial Resistance, Department of Microbiology and Infection Control, University Hospital of North-Norway, Tromsø, Norway
                [3 ] Department of Clinical Microbiology, Oslo University Hospital, Oslo, Norway
                [4 ] Institute of Clinical Medicine, University of Oslo, Oslo, Norway
                [5 ] Department of Medical Microbiology, St Olavs Hospital, Trondheim, Norway
                [6 ] Division of Nephrology, Department of Medicine, St Olavs Hospital, Trondheim, Norway
                Hospital Universitario Ramon y Cajal, SPAIN
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                https://orcid.org/0000-0001-9991-6315
                https://orcid.org/0000-0002-1314-0497
                Article
                PONE-D-21-10448
                10.1371/journal.pone.0255187
                8301612
                34297779
                22bef7a2-b2d3-43da-be7b-d0140b23cc60
                © 2021 AL Rubaye et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 3 May 2021
                : 11 July 2021
                Page count
                Figures: 3, Tables: 3, Pages: 15
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/501100007137, Helse Nord RHF;
                Award ID: HNF1362-17
                Award Recipient :
                Funded by: uit the arctic university of norway
                This work was supported by a grant from the Northern Norway Regional Health Authority Medical Research Programme (HNF1362-17). The publication charges for this article have been funded by a grant from the publication fund of UiT The Arctic University of Norway. The funders had no role in the study design, data collection and analysis, decision to publish or preparation of the manuscript.
                Categories
                Research Article
                Biology and Life Sciences
                Genetics
                Genomics
                Medicine and Health Sciences
                Pharmacology
                Drugs
                Antimicrobials
                Antibiotics
                Vancomycin
                Biology and Life Sciences
                Microbiology
                Microbial Control
                Antimicrobials
                Antibiotics
                Vancomycin
                Biology and Life Sciences
                Evolutionary Biology
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Biology and Life Sciences
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Computer and Information Sciences
                Data Management
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Biology and Life Sciences
                Microbiology
                Microbial Control
                Antimicrobial Resistance
                Antibiotic Resistance
                Vancomycin Resistance
                Medicine and Health Sciences
                Pharmacology
                Antimicrobial Resistance
                Antibiotic Resistance
                Vancomycin Resistance
                Biology and Life Sciences
                Organisms
                Bacteria
                Enterococcus
                Enterococcus Faecalis
                Biology and Life Sciences
                Microbiology
                Medical Microbiology
                Microbial Pathogens
                Bacterial Pathogens
                Enterococcus
                Enterococcus Faecalis
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Pathogens
                Microbial Pathogens
                Bacterial Pathogens
                Enterococcus
                Enterococcus Faecalis
                Biology and Life Sciences
                Organisms
                Bacteria
                Enterococcus
                Biology and Life Sciences
                Microbiology
                Medical Microbiology
                Microbial Pathogens
                Bacterial Pathogens
                Enterococcus
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Pathogens
                Microbial Pathogens
                Bacterial Pathogens
                Enterococcus
                Biology and Life Sciences
                Evolutionary Biology
                Evolutionary Systematics
                Phylogenetics
                Biology and Life Sciences
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Computer and Information Sciences
                Data Management
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Biology and Life Sciences
                Genetics
                Single Nucleotide Polymorphisms
                Custom metadata
                Genome raw reads and assemblies as well as the sequences of GIs generated in this study have been submitted to the NCBI. Raw reads have been deposited to the short read archive under Bioproject ID PRJNA627463. Genomic assemblies and island sequences have been deposited to NCBI under the following accession numbers: Genome sequences: JABBNS000000000, JABBNP000000000, JABBNR000000000, JABBNQ000000000, JABBNO000000000, and JABBNN000000000. GIs: MT951615, MT951616, and MT951617.

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